Aggrecan is a major extracellular component of articular cartilage. It is a proteoglycan responsible for providing cartilage with its mechanical properties of compressibility and elasticity. The loss of aggrecan has been implicated in the degradation of articular cartilage in arthritic diseases. Osteoarthritis is a debilitating disease which affects at least 30 million Americans (MacLean et al., J Rheumatol 25:2213–8 (1998)). Osteoarthritis can severely reduce quality of life due to degradation of articular cartilage and the resulting chronic pain. An early and important characteristic of the osteoarthritic process is loss of aggrecan from the extracellular matrix (Brandt and Mankin, Pathogenesis of Osteoarthritis, in Textbook of Rheumatology, WB Saunders Company, Philadelphia, Pa., at 1355–1373 (1993)). The large, sugar-containing portion of aggrecan is thereby lost from the extra-cellular matrix, resulting in deficiencies in the biomechanical characteristics of the cartilage.
A proteolytic activity termed “aggrecanase” is believed to be responsible for the cleavage of aggrecan thereby having a role in cartilage degradation associated with osteoarthritis and inflammatory joint disease. Research has been conducted to identify the enzymes responsible for the degradation of aggrecan in human osteoarthritic cartilage. At least two enzymatic cleavage sites have been identified within the interglobular domain of aggrecan. One enzymatic cleavage site within the interglobular domain of aggrecan (Asn341-Phe342) has been observed to be cleaved by several known metalloproteases. Flannery et al., J Biol Chem 267:1008–14 (1992); Fosang et al., Biochemical J. 304:347–351 (1994). Cleavage at a second aggrecan cleavage site within aggrecan (Glu373-Ala374) due to IL-1 induced cartilage aggrecan cleavage results in the generation of an aggrecan fragment found in human synovial fluid (Sandy et al., J Clin Invest 69:1512–1516 (1992); Lohmander et al., Arthritis Rheum 36: 1214–1222 (1993); Sandy et al., J Biol Chem 266: 8683–8685 (1991)). Aggrecan cleavage at (Glu373-Ala374) has been attributed to aggrecanase activity (Sandy et al., J Clin Invest 69:1512–1516 (1992). This Glu373-Ala374 cleavage site will be referred to as the aggrecanase cleavage site.
Recently, identification of two enzymes, aggrecanase-1 (ADAMTS4) and aggrecanase-2 (ADAMTS-11) within the “Disintegrin-like and Metalloprotease with Thrombospondin type 1 motif” (ADAMTS) family have been identified which are synthesized by IL-1 stimulated cartilage and cleave aggrecan at the Glu373-Ala374 site (Tortorella et al., Science 284:1664–6 (1999); Abbaszade et al., J Biol Chem 274: 23443–23450 (1999)). It is possible that these enzymes could be synthesized by osteoarthritic human articular cartilage. It is also contemplated that there are other, related enzymes in the ADAMTS family which are capable of cleaving aggrecan at the Glu373-Ala374 bond and could contribute to aggrecan cleavage in osteoarthritis. Therefore, there is a need to identify various aggrecanase enzymes and determine ways to block their enzymatic activity.